Exploring the Gastrointestinal “Nemabiome”: Deep Amplicon Sequencing to Quantify the Species Composition of Parasitic Nematode Communities

Parasitic helminth infections have a considerable impact on global human health as well as animal welfare and production. Although co-infection with multiple parasite species within a host is common, there is a dearth of tools with which to study the composition of these complex parasite communities. Helminth species vary in their pathogenicity, epidemiology and drug sensitivity and the interactions that occur between co-infecting species and their hosts are poorly understood. We describe the first application of deep amplicon sequencing to study parasitic nematode communities as well as introduce the concept of the gastro-intestinal “nemabiome”. The approach is analogous to 16S rDNA deep sequencing used to explore microbial communities, but utilizes the nematode ITS-2 rDNA locus instead. Gastro-intestinal parasites of cattle were used to develop the concept, as this host has many well-defined gastro-intestinal nematode species that commonly occur as complex co-infections. Further, the availability of pure mono-parasite populations from experimentally infected cattle allowed us to prepare mock parasite communities to determine, and correct for, species representation biases in the sequence data. We demonstrate that, once these biases have been corrected, accurate relative quantitation of gastro-intestinal parasitic nematode communities in cattle fecal samples can be achieved. We have validated the accuracy of the method applied to field-samples by comparing the results of detailed morphological examination of L3 larvae populations with those of the sequencing assay. The results illustrate the insights that can be gained into the species composition of parasite communities, using grazing cattle in the mid-west USA as an example. However, both the technical approach and the concept of the ‘nemabiome’ have a wide range of potential applications in human and veterinary medicine. These include investigations of host-parasite and parasite-parasite interactions during co-infection, parasite epidemiology, parasite ecology and the response of parasite populations to both drug treatments and control programs.     

Tracking Public Beliefs About Anthropogenic Climate Change

A simple question about climate change, with one choice designed to match consensus statements by scientists, was asked on 35 US nationwide, single-state or regional surveys from 2010 to 2015. Analysis of these data (over 28,000 interviews) yields robust and exceptionally well replicated findings on public beliefs about anthropogenic climate change, including regional variations, change over time, demographic bases, and the interacting effects of respondent education and political views. We find that more than half of the US public accepts the scientific consensus that climate change is happening now, caused mainly by human activities. A sizable, politically opposite minority (about 30 to 40%) concede the fact of climate change, but believe it has mainly natural causes. Few (about 10 to 15%) say they believe climate is not changing, or express no opinion. The overall proportions appear relatively stable nationwide, but exhibit place-to-place variations. Detailed analysis of 21 consecutive surveys within one fairly representative state (New Hampshire) finds a mild but statistically significant rise in agreement with the scientific consensus over 2010–2015. Effects from daily temperature are detectable but minor. Hurricane Sandy, which brushed New Hampshire but caused no disaster there, shows no lasting impact on that state’s time series—suggesting that non-immediate weather disasters have limited effects. In all datasets political orientation dominates among individual-level predictors of climate beliefs, moderating the otherwise positive effects from education. Acceptance of anthropogenic climate change rises with education among Democrats and Independents, but not so among Republicans. The continuing series of surveys provides a baseline for tracking how future scientific, political, socioeconomic or climate developments impact public acceptance of the scientific consensus.     

Genome‐wide analysis of Italian sheep diversity reveals a strong geographic pattern and cryptic relationships between breeds

Italy counts several sheep breeds, arisen over centuries as a consequence of ancient and recent genetic and demographic events. To finely reconstruct genetic structure and relationships between Italian sheep, 496 subjects from 19 breeds were typed at 50K single nucleotide polymorphism loci. A subset of foreign breeds from the Sheep HapMap dataset was also included in the analyses. Genetic distances (as visualized either in a network or in a multidimensional scaling analysis of identical by state distances) closely reflected geographic proximity between breeds, with a clear north–south gradient, likely because of high levels of past gene flow and admixture all along the peninsula. Sardinian breeds diverged more from other breeds, a probable consequence of the combined effect of ancient sporadic introgression of feral mouflon and long‐lasting genetic isolation from continental sheep populations. The study allowed the detection of previously undocumented episodes of recent introgression (Delle Langhe into the endangered Altamurana breed) as well as signatures of known, or claimed, historical introgression (Merino into Sopravissana and Gentile di Puglia; Bergamasca into Fabrianese, Appenninica and, to a lesser extent, Leccese). Arguments that would question, from a genomic point of view, the current breed classification of Bergamasca and Biellese into two separate breeds are presented. Finally, a role for traditional transhumance practices in shaping the genetic makeup of Alpine sheep breeds is proposed. The study represents the first exhaustive analysis of Italian sheep diversity in an European context, and it bridges the gap in the previous HapMap panel between Western Mediterranean and Swiss breeds.     

T-dependent destruction of thyroid isografts exposed to IFN-gamma

Several autoimmune diseases are accompanied by tissue-specific expression of class II molecules of the MHC, and it has been suggested that this elicits a T cell response against tissue-specific Ag to which the individual is not tolerant. However, recent transgenic studies have indicated that non-lymphoid expression of class II genes in the pancreas, liver, and kidney is either innocuous or induces peripheral tolerance. To test this hypothesis in another organ-specific autoimmune disease, we attempted to induce autoimmune thyroiditis in normal mice with class II+ thyroid tissue. Normal thyroid lobes were cultured with and without IFN-gamma and then transplanted to adult isogeneic recipients. The thyroid that had been induced to express class II genes by IFN-gamma was destroyed in normal mice, whereas the control cultured thyroid and the native cervical gland survived. Both types of transplants remained intact and functional in congenic nu/nu recipients, indicating that neither exposure to IFN-gamma nor expression of class II genes compromised the thyroid. Thus, in some tissues, exposure to IFN-gamma and/or the induction of class II expression can lead to T-dependent autoimmune disease.     

Regional distribution of blood flow within the diaphragm

We investigated the regional distribution of blood flow (Q) within the costal and crural portions of the diaphragm in a total of eight anesthetized supine mongrel dogs. Q was measured with 15-microns microspheres, radiolabeled with three different isotopes, injected into the left ventricle during spontaneous breathing (SB), inspiratory resistive loading (IR), and mechanical ventilation after paralysis (P). At necropsy, the costal and crural portions of each hemidiaphragm were arbitrarily subdivided along a sagittal plane into five to seven and three sections, respectively. During P, there was a dorsoventral Q gradient within the costal part of the diaphragm. During SB there was a fourfold increase in the gradient of Q. Furthermore, during IR, in which mouth pressures of -16 +/- 4 cmH2O were generated, there was a further increase in the gradient of Q. During both SB and IR, Q to the most ventral portion of the costal diaphragm was 26 +/- 6% less than the peak value. In two dogs, studied prone and supine, there was no difference in the Q gradients between the two postures. Over the dorsal 80% of the costal diaphragm there was also a dorsoventral gradient of muscle thickness, such that the most dorsal part was 54 +/- 2% (n = 5) that of the ventral portion. In contrast, there was no consistent gradient of Q or muscle thickness within the crural diaphragm. Our results demonstrate a topographical gravity-independent distribution of Q in the costal, but not the crural, diaphragm.(ABSTRACT TRUNCATED AT 250 WORDS)     

Iron-sulfur centers in the Staphylococcus aureus respiratory chain]

Using low temperature EPR spectroscopy, signals of iron-sulfur centers with g-factors of 2.02 and 1.94 were detected in the respiratory chain of St. aureus membranes. According to their relaxation parameters and redox properties, these iron-sulfur centers are similar to iron-sulfur centers S-1 and S-3 corresponding to succinate dehydrogenases of mitochondria and bacterial membranes.     

Two forms of sensitization of the local bending reflex of the medicinal leech

Sensitization of the local bending reflex of the medicinal leech Hirudo medicinalis was studied in a semi-intact preparation in which behavioral and electrophysiological recordings were made simultaneously. 1. Sensitization of local bending could be produced in two ways: by repeated stimulation of the mechanoreceptor sensitive to pressure (the P cell), and by stimulation of the mechanoreceptor sensitive to noxious stimuli (the N cell). 2. Both forms of sensitization produced a central neuronal change, measured as an increase in the number of stimulus-evoked action potentials in cell 3 (an excitor of dorsal longitudinal muscles). 3. Intracellular stimulation of serotonin-containing neurons 21 and 61 mimicked the sensitizing stimuli, but stimulation of the Retzius cell, which also contains serotonin, did not. 4. Stimulation of the Leydig cell, which releases octopamine, decreased the strength of local bending.     

Structural similarity and distribution of small cryptic plasmids of Lactobacillus curvatus and L. sake

Plasmid profiles of strains of Lactobacillus curvatus and L. sake isolated from meat or sauerkraut were analysed to investigate plasmid homology and distribution in relation to the ecology of these organisms in fermenting foods. A hybridisation probe was constructed by cloning of pLc2, a cryptic, 2.6-kbp plasmid from L. curvatus LTH683, into the Escherichia coli plasmid pRV50. In Southern hybridisations with the digoxygenine labeled pLc2 probe, pLc2-related small plasmids were frequently detected in meat-borne strains of L. casei subsp. pseudoplantarum, L. curvatus, L. sake, L. alimentarius, L. farciminis and L. halotolerans and in L. curvatus and L. sake isolated from sauerkraut. Among 27 Lactobacillus type strains originally isolated from habitats other than meat this type of homology was detected only with plasmids of L. buchneri and L. mali. Restriction-enzyme mapping of six small cryptic plasmids from L. curvatus and L. sake revealed strong structural homology but no similarity to previously characterized plasmids of lactobacilli. The presence of a variable region in addition to a conserved one and the occurrence of deletions during cloning of pLc2 suggest that vectors derived from these plasmids are likely to be structurally unstable.     

An early Holocene reef in the western Atlantic: submersible investigations of a deep relict reef off the west coast of Barbados,W.I.

Submersible observations and collections reveal that a probable relict reef off the west coast of Barbados has a rich cover of sponges, along with algae and scattered corals, on a substrate of algal nodules in a muddy-sand matrix. The collections provide new data on the distributions of these fauna. This relict reef is about 20 km long, has a relief of up to 10 m, and is established at a depth of 80 m. Relict shallow-water features in other areas at similar depths along with data from core holes drilled off the south coast of Barbados suggest that this reef was probably established about 12,000 years ago and existed for no more than 2,000 years, during the Holocene sea-level transgression.